针对结核分枝杆菌潜伏感染的DNA疫苗V569免疫原性及保护性的初步研究

Immunogenicity and protective results of DNA vaccine V569 against latent tuberculosis infection in animal models

为研究针对结核分枝杆菌潜伏感染的DNA疫苗,基于质粒A39构建了p-VAX1-Ag85B-Rv3425-Rv2029c-PPE26(V569)质粒DNA,并对其免疫原性及保护性进行初步研究。免疫性评价试验共分6组:PBS、p-VAX1-Ag85B(A)、p-VAX1-Ag85B-Rv3425(A3)、A39、V569和BCG,采用左后腿肌内注射C57BL/6小鼠,用流式细胞术和酶联免疫吸附试验(enzyme linked immunosorbent assay,ELISA)分别检测细胞免疫和体液免疫水平;构建斑马鱼-海分枝杆菌潜伏感染模型,将PBS、A、A3、A39、BCG、V569分别通过腹腔注射免疫斑马鱼后,每日注射地塞米松10ug诱导海分枝杆菌复发感染,对斑马鱼肝脏进行菌落计数并绘制生存曲线。结果显示,与BCG组相比,V569能引发实验小鼠强烈的细胞免疫反应(IFN-γ高水平分泌),外周血CD4+/CD8+T细胞比例明显增加。在斑马鱼-海分枝杆菌潜伏感染复发模型中,与BCG免疫组相比,V569免疫斑马鱼后可显著减少其肝脏中海分枝杆菌数量,斑马鱼存活情况得到显著改善,表明V569 DNA疫苗可能是一种抗结核潜伏感染的候选DNA疫苗。

We constructed the DNA vaccine Ag85B-Rv3425-Rv2029c-PPE26(V569)plasmid based on p-VAX1-Ag85B-Rv3425-Rv2029c(A39)and evaluated its immunogenicity in mice.The levels of cellular immunity and humoral immunity against PBS,BCG,p-VAX1-Ag85B(A),p-VAX1-Ag85B-Rv3425(A3),A39 and V569 were detected by enzyme-linked immunosorbent assay(ELISA)and flow cytometry after vaccination in mice.We found that V569 could trigger stronger Th1 immune response by augmenting the secretion of IFN-γand significantly improve the CD4+/CD8+T cell ratio in immunized mice compared to BCG.In addition,we evaluated the V569 as post-exposure DNA vaccine and found that it reduced bacterial burdens upon reactivation compared to BCG.The protection of V569 against latent TB infection was determined by zebrafish-Mycobacterium marinum latent infection model.In conclusion,V569 DNA vaccine may be a potential candidate DNA vaccine against latent tuberculosis infection.